The major goal of this project is to understand, at the cell and molecular level, the functions of the novel transcription factor RFX4_v3 in brain development. The RFX4_v3 transcription factor was originally described by our group in transgenic insertional mutagenesis studies as a factor that, when completely deficient, led to grossly abnormal brain development following the complete failure to form of brain midline structures. The single allelic deficiency mice developed non-communicating hydrocephalus, with near complete failure of development of the subcommissural organ, the structure that controls the secretion of Reissner's fibers that in turn are necessary for patency of the aqueduct of Silvius. Subsequent studies focused on the potential promoter targets for this transcription factor and associated proteins as determined by two hybrid experiments. We recently succeeded in developing a conditional allele for this locus. To date, the analysis of mice with a "total body" deletion of this locus has revealed very similar phenotypes to those described above for single and double allelic knockouts using the transgene inertional mutagenesis technique. Our ongoing studies plan to use these conditional KO mice to understand the role of this trancription factor in the development of the central nervous system by doing selective removal in different cell lineages and during different stages of development. Finally, we are interested in eventually determining whether naturally occurring variants or polymorphisms in the genomes of human populations lead to predispositions or increased susceptibility to the develoment of congenital central nervous system defects or congenital hydrocephalus, perhaps by interacting with one or multiple environmental influences. These studies have been being expanded into the evaluation of the promoter activity that leads to expression of this transcription factor. Essentially nothing is known about the regulation of this promoter, which is remarkably conserved among all vertebrate species examined to date, and which contains a number of transcription factor binding sites known to be important for neural development.